分生实验（Meristem experiment）.docVIP

分生实验（Meristem experiment） 1. what is the function of the reagents used in this experiment? Answer: NE solution: metal ion chelating agent, inhibit the activity of DNase; Protease K: broad spectrum protease can maintain high activity in the presence of SDS and EDTA, degrade proteins and dissociate DNA; SDS:a. anionic detergent dissolves lipid components in cell membranes and nuclear membranes; B. denatured proteins; C. destroys cell membranes and nuclear membranes and inhibits nuclease; TE saturated phenols: impurities such as protein removal. Phenol has a strong denaturation of proteins, which can cause protein denaturation in the sample and be removed by centrifugation. In order to prevent its influence on the subsequent experiments, chloroform was usually extracted to remove residual phenols; Chloroform: the degeneration of chloroform of A. is better than that of phenol, but it can enhance the effect of removing protein if it is used in combination with phenol; B. chloroform accelerates the separation of organic and aqueous phases, removes plant pigments and sucrose, and inhibits nuclease activity; Removal of Trace Phenols in DNA aqueous solution by c.; Isoprene: reduce the formation of bubbles during operation; Ethanol: in the presence of monovalent cations, ethanol can precipitate DNA; Sodium acetate: provides monovalent cations; EDTA: inhibits the activity of DNA enzymes; What are the considerations in the 2.DNA extraction process? Answer: 1) this experiment will be used TE saturated, heavy steaming phenol, chloroform / isoamyl alcohol is organic reagent, than water gravity, and DNA dissolved in water, we always take supernatant. 2) in the extraction of phenol, chloroform and other organic reagents, you can not put the sampler flat or upside down, in order to prevent liquid flow back damage to the sampler, after the sample, immediately discard the sampler head. 3) genome DNA is too large to break down by mechanical shearing, so try not to do physical operations